Anti-Retroviral Therapy (ART) targets HIV-1 replication in CD4+ T cells that are active in virus production. Although ART can reduce the viral load to undetectable levels, a small population of drug-resistant viral genomes is selected and continues to replicate in the presence of the drugs. In this population, the molecular targets of antiretroviral therapy, reverse transcriptase (RT) and protease (PR), accumulate mutations that confer drug resistance. At the same time these drug-resistance mutations are deleterious, reducing the activity of RT and PR and resulting in the persistence of viruses that have lower replicative capacity (i.e., lesser "fitness"). Continuing viral evolution selects for compensating mutations resulting in the emergence of viruses that are fitter. Such mutations in the RT and PR genes are well characterized, but there are also indications that compensating mutations can arise in other viral genes. One such candidate gene is the essential viral transcription activator, Tat. We and others have identified natural Tat mutations that increase viral gene expression. We postulate that viruses with mutations in Tat, or in its viral target TAR, increase the fitness of multidrug-resistant viruses and will be selected for survival. In this exploratory project we propose to address the hypothesis that Tat and/or TAR variants of higher potency are selected during ART. We will test the fitness of viruses containing these selected Tat and/or TAR variants in the context of drug-resistance mutations in RT and PR, and subsequent experiments will address the underlying biochemical mechanisms. During the term of this grant our aims will be: 1. To sequence and analyze HIV-1 Tat genes and the TAR element of viruses isolated from HIV-1 drug-resistant patients. 2. To characterize the replicative fitness and function of selected Tat and TAR variants.